Thrombin, a coagulation protease generated at sites of vascular injury, activates platelets, leukocytes, and mesenchymal cells (Vu, T. -K. H. et al. (1991) Cell 64:1057–1068). Activation of platelets by thrombin is thought to be critical for hemostasis and thrombosis. In animal models, thrombin inhibitors block platelet-dependent thrombosis, which is the cause of most heart attacks and strokes in humans. Available data in humans suggests that thrombosis in arteries can be blocked by inhibitors of platelet function and by thrombin inhibitors. Thus it is likely that thrombin's actions on platelets contribute to the formation of clots that cause heart attack and stroke. Thrombin's other actions on vascular endothelial cells and smooth muscle cells, leukocytes, and fibroblasts may mediate inflammatory and proliferative responses to injury, as occur in normal wound healing and a variety of diseases (atherosclerosis, restenosis, pulmonary inflammation (ARDS), glomerulosclerosis, etc.). A thorough understanding of how thrombin activates cells is an important goal.
A receptor that mediates thrombin signaling has been previously identified (Vu, T. -K. H. et al. (1991) Cell 64:1057–1068; U.S. Pat. No. 5,256,766). This receptor revealed a novel proteolytic mechanism of activation and is referred to as PAR1 (protease-activated receptor 1). PAR1 is activated by the binding of thrombin to and cleavage of PAR1's amino terminal exodomain at a specific site. Receptor cleavage unmasks a new amino terminus, which then functions as a tethered peptide ligand by binding intramolecularly to the body of the receptor to effect transmembrane signaling (Vu, T. -K. H. et al. (1991) Cell 64:1057–1068). Synthetic peptides that mimic this tethered ligand domain function as PAR1 agonists and activate it independent of thrombin and receptor cleavage (Vu, T. -K. H. et al. (1991) Cell 64:1057–1068).
To identify which of thrombin's known cellular actions are mediated by PAR1, a PAR1 knockout mouse was recently generated (Connolly, A. et al. (1996) Nature 381:516–519). Analysis of mice in which both alleles of the PAR1 gene were disrupted provided definitive evidence for a second platelet thrombin receptor and for tissue specific roles of distinct thrombin receptors. Specifically, in mice, PAR1 is not important for platelet responses but is critical for fibroblast responses.
A second protease-activated receptor (PAR2) was cloned during a search for relatives of the Substance K receptor (Nystedt, S., et al. (1994) PNAS USA, 91:9208–9212). The physiological activator of PAR2 remains unknown; it is not activated by thrombin.